3,6-hemiketals from the class of 9A-azalides

ABSTRACT

The invention relates to novel compounds from the class of macrolide antibiotics. Particularly, the invention relates to novel 3,6-heiketals from the class of 9a-azalides, to their pharmaceutically acceptable addition salts with inorganic or organic acids, to a process for their preparation and to the use thereof as antibiotics or as intermediates for the synthesis of other macrolide antibiotics.

TECHNICAL FIELD OF THE INVENTION

A61 K 31/70, C 07 H 17/08

TECHNICAL PROBLEM

The invention relates to novel compounds from the class of macrolideantibiotics. Particularly, the invention relates to novel 3,6-hemiketalsfrom the class of 9a-azalides, to their pharmaceutically acceptableaddition salts with inorganic or organic acids, to a process for theirpreparation and to the use thereof as antibiotics or as intermediatesfor the synthesis of other macrolide antibiotics.

PRIOR ART

Macrolide antibiotic erythromycin A has been for more than 40 yearsconsidered as a safe and efficient agent for the treatment ofrespiratory and genital infections caused by Gram-positive and by someGram-negative bacteria, some species of Legionella, Mycoplasma,Chlamidia and Helicobacter. Noticed changes in bioavailability afteroral administration, gastric intolerance in many patients and loss ofactivity in an acidic medium whereat the inactive metaboliteanbydroerythromycin is formed are basic disadvantages in the clinicaluse of eiythromycin. However, the spirocyclization of the aglycone ringis successfully inhibited by a chemical transformation of C-9 ketone orhydroxyl groups in C-6 and/or C-12 positions. Thus, e.g by oximation ofC-9 ketone and subsequent Beckmann rearrangement and reduction,9-deoxo-9a-aza-9a-homoerythromycin A, the first 15-membered macrolideantibiotic with 9a-amino group incorporated in the aglycone ring, isobtained (Kobrehel G. et al., U.S. Pat. No. 4,328,334; May 1982). Byreductive methylation of 9-amines according to Eschweiler-Clark process,9-deoxo-9a-methyl-9a-aza-9a-homoerythromycin (AZITHROMYCIN), a prototypeof a novel class of macrolide antibiotics, namely azalides, issynthesized (Kobrehel G. et al., BE 892357; July 1982). In addition to abroad antimicrobial spectrum including also Gram-negative bacteria,azithromycin is also characterized by a long biological half-life, aspecific transport mechanism to the place of use and a short therapyperiod. Azithromycin easily penetrates and it accumulates inside humanphagocyte cells resulting in an improved action upon intracellularpathogenic micro-organisms from the classes of Legionella, Chlamidia andHelicobacter.

Further, it is known that C-6/C-12 spirocyclization of erythromycin A issuccessfully inhibited by O-methylation of C-6 hydroxyl group of theaglycone ring (Watanabe Y. et al., U.S. Pat. No. 4,331,803; May 1982).By the reaction of erythromycin with benzyloxycarbonyl chloride andsubsequent methylation of the obtained 2′-O,3′-N-bis(benzyloxycarbonyl)derivative, by elimination of the protecting groups and by3′-N-methylation, there are formed, in addition to6-O-methylerythromycin (CLARITHROMYCIN), also significant amounts of11-O-methylerythromycin and of multiple-substituted analogs (MorimotoS., et al., J. Antibiotics, 1984, 37, 187). With respect to erytlromycinA, clarithromycin is considerably more stable in an acidic medium andexhibits better in vitro action with respect to Gram-positive bacteriastrains (Kirst H. A. et al., Antimicrobial Agents and Chemoter., 1989,1419). In a similar manner also a series of O-methyl-derivatives ofazithromycin (Kobrehel G. et al., U.S. Pat. No. 5,250,518; October 1993)was synthesized. Although the main products of O-methylation ofazithromycin, namely 11-O-methyl-azithromycin (Example 8) and6-O-methyl-azithromycin (Example 6) exhibit significant activity againststandard bacteria strains and clinical isolates and pharmacokineticproperties similar to those of azithromycin, the obtaining of productsin larger quantities represents an additional technical problem due tononselectivity of O-methylation. The determination of the structure ofO-methyl-derivatives of azithromycin was based on analysis of ¹H-¹H and¹H-¹³C 2D NMR spectra (300 MHz). Subsequently, it was additionallydetermined by long-range NMR spectroscopy that substitution on C-6hydroxyl group had been erroneously ascribed to azithromycin and thatactually 12-O-methyl-azithromycin was in question. Further it has beenfound that the use of suitable protecting groups on hydroxyl groups in4″- and 11-positions (especially of silyl protecting groups such astrimethylsilyl groups) results in selective O-methylation and makespossible a simple preparation of 12-O-methyl-azithromycin (HR 970051A;October 1997). Later, Waddell S. T. et al., (Biorg. Med. Chem. Letters 8(1998), 549-555), independently of the latter patent application,established O-methylation of hydroxyl group in C-12 position.

It is known as well that recent research on 14-membered macrolides haslead to the discovery of a new type of macrolide antibiotics, namelyketolides. Instead of the neutral sugar L-cladinose known for itsunstability even in a weakly acidic medium, these compounds possess aketo group on C-3 position (Agouridas C. et al., EP 596802 A1, May 1994;Le Martret O., FR 2697524 A1, May 1994). Ketolides show a significantlybetter action against MLS (macrolide, lincosamide and streptogramin B)induced-resistant organisms (Jamjian C., Antimicrob. Agents Chemother.,1997, 41, 485). This important discovery has led to a large number of3-keto derivatives of clarithromycin, mostly substituted on C-11/C-12positions, yielding numerous cyclic carbonates, carbamates and,recently, carbazates. The first step of the synthesis of ketolidesincludes the hydrolysis of clarithromycin under the formation of acorresponding 3-decladinosyl derivative,(3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-derivative),which is, after the removal of the protection of 2′-hydroxyl group(preferably by acylation with chlorides or anhydrides of carboxylicacids), subjected to a reaction of oxidation and deprotection of 2′-position. According to our knowledge C-11/C-12 substituted ketolidesfrom the class of 9a-azalide antibiotics have hitherto not beendescribed. The first step, namely the synthesis of3-decladinosyl-derivatives of 9-deoxo-9a-aza-9a-homoerydiromycin andazithromycin, is described in U.S. Pat. No. 4,886,792, December 1989.With intention to oxidize C-3 hydroxyl group of3-decladinosyl-azithromycin and its 11-O-methyl- and12-O-methyl-derivatives by transannular addition of 6-hydroxyl grouponto the newly formed C-3 ketone there has been obtained a hitherto notdescribed series of bicyclic and tricyclic 3,6-hemiketals from the classof 9a-azalides.

The synthesis of 3,6-hemiketals of azithromycin and O-methyl derivativesthereof comprises the preparation of corresponding 3-decladinosylderivatives, the protection of 2′-hydroxyl group of the basic sugar,D-desosamine, by selective acylation, the oxidation of the hydroxylgroup in C-3 position, the deprotection of 2′-position and thecyclization of C-11 and C-12 hydroxyl groups. Objects of the presentinvention are also pharmaceutically acceptable addition salts of3,6-hemiketals of azithromycin and its O-methyl derivatives with organicand inorganic acids, methods and intermediates for their preparation, aswell as preparation and application methods of pharmaceuticalpreparations.

DESCRIPTION OF TECHNICAL PROBLEM WITH EXAMPLES

The invention relates to

i) novel 3,6-hemiketals from the class of 9a-azalides,

ii) a process for the preparation of novel 3,6-hemiketals from the classof 9a-azalides.

iii) use of novel 3,6-hemiketals from the class of 9a-azalides asantibiotics or as intermediates for the synthesis of other macrolideantibiotics.

Novel 3,6-hemiketals from the class of 9a-azalides of the generalformula (I)

characterized in that

R¹ individually stands for hydroxyl, L-cladinosyl group of the formula(II)

wherein

R² individually stands for hydrogen or a silyl group,

R³ individually stands for hydrogen or together with R⁶ stands for anether group,

R⁴ individually stands for hydrogen, (C₁-C₄)acyl group or—COO—(CH₂)_(n)—Ar group, wherein n is 1-7 and Ar individually stands foran unsubstituted or substituted aryl group with up to 18 carbon atoms,

R⁵ individually stands for hydrogen, methyl group or —COO—(CH₂)_(n)—Argroup, wherein n is 1-7 and Ar individually stands for an unsubstitutedor substituted aryl group with up to 18 carbon atoms,

R⁶ individually stands for a hydroxyl group or together with R³ has themeaning of an ether group,

R⁷ individually stands for hydrogen, (C₁-C₁₂)alkyl group, silyl group ortogether with R⁸ and C-11/C-12 carbon atoms stands for a cycliccarbonate,

R⁸ individually stands for hydrogen, (C₁-C₁₂)alkyl group, silyl group ortogether with R⁷ and C-11/C-12 carbon atoms stands for a cycliccarbonate,

and their pharmaceutically acceptable addition salts with inorganic ororganic acids, are obtained by the following steps.

Step 1

Azithromycin of the general formula (I) wherein R¹ stands forL-cladinosyl group of the formula (II), R², R³, R⁴, R⁷ and R⁸ aremutually the same and stand for hydrogen, R⁵ is methyl and R⁶ is ahydroxyl group, is subjected to a reaction with organic carboxylic acidchlorides of the formula (III)

 ClCOO(CH₂)_(n)—Ar  (III)

wherein n is 1-7 and Ar individually stands for unsubstituted orsubstituted aryl groups with up to 18 carbon atoms, preferably withbenzyloxycarbonyl chloride, in the presence of bases, preferably sodiumhydrogen carbonate, in a reaction-inert solvent, preferably in benzeneor toluene, yielding 2′-O,3′-N-bis(bezyloxycarbonyl)-3′(Kobrehel G. etal., U.S. Pat. No. 5,250,518; May 1993) of the general formula (I),wherein R¹ stands for L-cladinosyl group of the formula (II), R², R³, R⁷and R⁸ are mutually the same and stand for hydrogen, R⁴ and R⁵ aremutually the same and stand for benzyloxycarbonyl group and R⁶ ishydroxyl group, which is subsequently subjected to silylation ofhydroxyl groups in

A/ 4″- and 11-positions with 2-5 equimolar excess of a silylating agent,in an organic inert solvent, at the temperature of 0-5° C. during 5-8hours, yielding novel4″-11-O-bis(trimethylsilyl)-2′-O,3′-N-bis(benzyloxycarbonyl)-3′-N-demethyl-azithromycinof the general formula (I), wherein R¹ stands for L-cladinosyl group ofthe formula (II), R² and R⁷ are mutually the same and stand fortrimethylsilyl group, R³ and R⁸ are mutually the same and stand forhydrogen, R⁴ and R⁵ are mutually the same and stand forbenzyloxycarbonyl group and R⁶ is hydroxyl group, or in

B/ 4″-position with 1.1-2 equimolar excess of a silylating agent, in anorganic inert solvent, at the temperature of 0-5° C. during 1 hour,yielding novel4″-O-trimethyl-silyl-2′-O,3′-N-bis(benzyloxycarbonyl)-3′-N-demethyl-azithromycinof the general formula (I), wherein R¹ stands for L-cladinosyl group ofthe formula (II), R² stands for trimethylsilyl group, R³, R⁷ and R⁸ aremutually the same and stand for hydrogen, R⁴ and R⁵ are mutually thesame and stand for benzyloxycarbonyl group and R⁶ stands for hydroxylgroup.

As silylating agents there are used 1,1,1,3,3,3-hexamethyldisilazane,trimethylsilyl chloride, bis(trimethylsilyl)acetamide and similar agentsfor introducing trimethylsilyl group, preferably a mixture oftrimethylsilyl chloride and trimethylsilyl imidazole. As a suitablesolvent pyridine, ethyl acetate, N,N-dimethylformamide, methylenechloride and the like, preferably pyridine are used.

Step 2

By a reaction of4″,11-O-bis(trimethylsilyl)-2′-O,3′-N-bis(benzyloxycarbonyl)-3′-N-demethyl-azithromycinfrom the step 1A/or4″-O-trimethylsilyl-2′-O,3′-N-bis(benzyl-oxycarbonyl)-3′-N-demethyl-azithromycinfrom the step 1B/, respectively, with 1.3-10 moles of a correspondingalkylating agent, preferably methylating agent, in the presence of1.1-8.5 moles of a suitable base, at a temperature from −15° C. to roomtemperature, preferably at 0-5° C., in a suitable reaction-inertsolvent, there comes to

A/ a selective alkylation, preferably methylation of C-12 hydroxyl groupyielding a novel4″-11-O-bis(trimethylsilyl)-2′-O,3′-N-bis(benzyloxycarbonyl)-3′-N-demethyl-12-O-methyl-azithromycinof the general formula (I), wherein R¹ stands for L-cladinosyl group ofthe formula (II), R² and R⁷ are mutually the same and stand fortrimethylsilyl group, R³ stands for hydrogen, R⁴ and R⁵ are mutually thesame and stand for benzyloxycarbonyl group, R⁶ is hydroxyl group and R⁸is methyl, or

B/ an alkylation, preferably methylation of C-11 or C-12 hydroxyl groupyielding a mixture of novel4″-O-trimethylsilyl-2′-O,3′-N-bis(benzyloxycarbonyl)-3′-N-demethyl-11-O-methyl-azithromycinof the general formula (I), wherein R¹ stands for L-cladinosyl group ofthe formula (II), R² stands for trimethylsilyl group, R³ and R⁸ aremutually the same and stand for hydrogen, R⁴ and R⁵ are mutually thesame and stand for benzyloxycarbonyl group, R⁶ stands for hydroxyl groupand R⁷ is methyl, or4″-O-trimethylsilyl-2′-O,3′-N-bis(benzyloxycarbonyl)-3′-N-demethyl-12-O-methyl-azithromycinof the general formula (I), wherein R¹ stands for L-cladinosyl group ofthe formula (II), R² stands for trimethylsilyl group, R³ and R⁷ aremutually the same and stand for hydrogen, R⁴ and R⁵ are mutually thesame and stand for benzyloxycarbonyl group, R⁶ stands for hydroxyl groupand R⁸ is methyl.

As suitable alkylating agents there are used (C₁-C₁₂)alkyl halides,preferably methyl iodide, dimethyl sulfate, methyl methane sulfonate ormethyl p-toluene sulfonate, preferably methyl iodide. Suitable bases arealkali metal hydride (lithium hydride, sodium hydride or potassiumhydride), alkali metal hydroxide (potassium hydroxide or sodiumhydroxide) or alkali metal methyl amide (lithium amide, sodium amide orpotassium amide), preferably sodium hydride. Suitable reaction-inertsolvents are dimethyl sulfoxide, N,N-dimethyl formamide, N,N-dimethylacetamide or hexamethyl phosphoric triamide, preferably N,N-dimethylformamide, dimethyl sulfoxide or a mixture thereof with tetrahydrofuran.

Step 3

4″-11-O-Bis(trimethylsilyl)-2′-O,3′-N-bis(benzyloxycarbonyl)-3′-N-demethyl-12-O-methyl-azithromycinfrom the step 2A/ or the obtained mixture of4″-O-trimethylsilyl-2′-O,3′-N-bis(benzyloxycarbonyl)-3′-N-demethyl-11-O-methyl-azithromycinand4″-O-trimethylsilyl-2′-O,3′-N-bis(benzyloxycarbonyl)-3′-N-demethyl-12-O-methyl-azithromycinfrom the step 2B/ is subjected to a hydrogenolysis reaction according tothe method by E. H. Flynn et al. (Journal of American Chemical Society,77, 3104, 1950) in order to deprotect protecting groups on 2′- and3′-positions and then to desilylation according to the conventionalprocess in lower alcohols, preferably isopropanol in the presence offormic acid in

A/ 4″- and 11-positions in the step 2A/ yielding3′-N-demethyl-12-O-methyl-azithromycin of the general formula (I)wherein R¹ stands for L-cladinosyl group of the formula (II), R², R³,R⁴, R⁵ and R⁷ are mutually the same and stand for hydrogen, R⁶ ishydroxyl group and R⁸ is methyl, or in

B/ 4″-position in the Step 2B/ yielding a mixture of3′-N-demethyl-11-O-methyl-azithromycin of the general formula (I),wherein R¹ stands for L-cladinosyl group of the formula (II), R², R³,R⁴, R⁵ and R⁸ are mutually the same and stand for hydrogen, R⁶ ishydroxyl group and R⁷ is methyl, and3′-N-demethyl-12-O-methyl-azithromycin of the general formula (I),wherein R¹ stands for L-cladinosyl group of the formula (II), R², R³,R⁴, R⁵ and R⁷ are mutually the same and stand for hydrogen, R⁶ ishydroxyl group and R⁸ is methyl.

Hydrogenolysis is carried out in a solution of lower alcohols,preferably in ethanol, in the presence of NaOAc/HOAc buffer (pH 5) witha catalyst such as palladium black or palladium on charcoal, at ahydrogen pressure from 1 to 20 bars, at room temperature.

Step 4

3′-N-Demethyl-12-O-methyl-azithromycin from the step 3A/ or the obtainedmixture of 3′-N-demethyl-11-O-methyl-azithromycin and 3 ′-N-demethyl-12-O-methyl-azithromycin from the Step 3B/ is subjected to a reductive3′-N-methylation with 1-3 equivalents of formaldehide (37%) in thepresence of an equal or double quantity of formic acid (98-100%) andhydrogenation catalyst or of some other hydrogen source, in areaction-inert solvent such as halogenated hydrocarbons, lower alcoholsor lower ketones, preferably chloroform, at the reflux temperature ofthe reaction mixture, yielding—in the case of the compound from the Step3A/—12-O-methyl-azithromycin of the general formula (I), wherein R¹stands for L-cladinosyl group of the formula (II), R², R³, R⁴ and R⁷ aremutually the same and stand for hydrogen, R⁵ and R⁸ are mutually thesame and stand for methyl and R⁶ is hydroxyl group, or—in the case ofproducts from the Step 3B/—a mixture of 11-methyl-azithromycin of thegeneral formula (I), wherein R¹ stands for L-cladinosyl group of theformula (II), R², R³, R⁴ and R⁸ are mutually the same and stand forhydrogen, R⁵ and R⁷ are mutually the same and stand for methyl and R⁶ ishydroxyl group, and of 12-O-methyl-azithromycin of the general formula(I), wherein R¹, R², R³, R⁴, R⁵, R⁶, R⁷ and R⁸ have the meanings asgiven in the case of 3′-N-methylation of the compounds from the Step3A/.

Step 5

Azithromycin of the general formula (I), wherein R¹ stands forL-cladinosyl group of the formula (II), R², R³, R⁴, R⁷ and R⁸ aremutually the same and stand for hydrogen, R⁵ is methyl and R⁶ ishydroxyl group, or its 11-O-methyl- and 12-O-methyl-derivatives from theStep 4 are optionally subjected to hydrolysis with strong acids,preferably with 0.25-1.5 N hydrochloric or dichloroacetic acid in amixture of water and an alcohol, preferably methanol, ethanol orisopropanol, for 10-30 hours, at room temperature yielding3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-azithromycinof the general formula (I), wherein R¹ and R⁶ are mutually the same andstand for hydroxyl group, R³, R⁴, R⁷ and R⁸ are mutually the same andstand for hydrogen and R⁵ is methyl, or3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-11-O-methyl-azithromycinof the general formula (I), wherein R¹ and R⁶ are mutually the same andstand for hydroxyl group, R³, R⁴ and R⁸ are mutually the same and standfor hydrogen and R⁵ and R⁷ are mutually the same and stand for methyl,or3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-12-O-methyl-azithromycinof the general formula (I), wherein R¹ and R⁶ are mutually the same andstand for hydroxyl group, R³, R⁴ and R⁷ are mutually the same and standfor hydrogen and R⁵ and R⁸ are mutually the same and stand for methyl.

Step 6

3-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-azithromycinand its 11-O-methyl- and 12-O-methyl derivatives from the Step 5 aresubjected to a selective acylation of the hydroxyl group in 2′-position.Acylation is carried out with chlorides or anhydrides of carboxylicacids with up to 4 carbon atoms, preferably with acetic acid anhydride,in the presence of inorganic or organic bases, in a reaction-inertorganic solvent, at a temperature from 0-30° C., yielding3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-azithromycin2′-O-acetate of the general formula (I), wherein R¹ and R⁶ are mutuallythe same and stand for hydroxid group, R³, R⁷ and R⁸ are mutually thesame and stand for hydrogen, R⁴ is acetyl and R⁵ is methyl, or3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-11-O-methyl-azithromycin2′-O-acetate of the general formula (I), wherein R¹ and R⁶ are mutuallythe same and stand for hydroxyl group, R³ and R⁸ are mutually the sameand stand for hydrogen, R⁴ is acetyl and R⁵ and R⁷ are mutually the sameand stand for methyl, or3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-12-O-methyl-azithromycin 2′-O-acetate of the general formula (I),wherein R¹ and R⁶ are mutually the same and stand for bydroxyl group, R³and R⁷ are mutually the same and stand for hydrogen, R⁴ is acetyl and R⁵and R⁸ are mutually the same and stand for methyl.

As suitable bases there are used sodium hydrogen carbonate, sodiumcarbonate, potassium carbonate, triethylamine, pyridine, tributylamine,preferably sodium hydrogen carbonate. As a suitable inert solvent thereis used methylene chloride, dichloroethane, acetone, pyridine, ethylacetate, tetrahydrofuran, preferably methylene chloride.

Step 7

3-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-azithromycin2′-O-acetate and its 11-O-methyl- and 12-O-methyl derivatives from theStep 6 are subjected to oxidation of the hydroxyl group in C-3 positionwith Jones reagent or diimides according to a modified Moffat-Pfitznerprocess [DMSO and 1-(3-dimethylaminopropyl)-3-ethyl-carbodiimide in thepresence of pyridine trifluoro-acetate] yielding3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-azithromycin3,6-hemiketal 2′-O-acetate of the general formula (I), wherein R¹ standsfor hydroxyl group, R³ together with R⁶ stands for an ether group, R⁴ isacetyl, R⁵ is methyl, and R⁷ and R⁸ are mutually the same and stand forhydrogen, or3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-11-O-methyl-azithromycin 3,6-hemiketal 2′-O-acetate of the generalformula (I), wherein R¹stands for hydroxyl group, R³ together with R⁶stands for an ether group, R⁴ is acetyl, R⁵ and R⁷ are mutually the sameand stand for methyl, and R⁸ is hydrogen, or3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribo-hexopyranosyl-oxy)-12-O-methyl-azithromycin3,6-hemiketal 2′-O-acetate of the general formula (I), wherein R¹ standsfor hydroxyl group, R³ together with R⁶ stands for an ether group, R⁴ isacetyl, R⁵ and R⁸ are mutually the same and stand for methyl and R⁷ ishydrogen.

Step 8:

3-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-azithromycin3,6-hemiketal 2′-O-acetate and its 11-O-methyl- and12-O-methyl-derivatives from the Step 7 are subjected to solvolysis inlower alcohols, preferably in methanol, at a temperature from roomtemperature to the reflux temperature of the solvent, yielding3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-azithromycin3,6-hemiketal of the general formula (I), wherein R¹ stands for hydroxylgroup, R³ together with R⁶ stands for an ether group, R⁴, R⁷ and R⁸ aremutually the same and stand for hydrogen, and R⁵ is methyl, or3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-11-O-methyl-azithromycin 3,6-hemiketal of the general formula (I),wherein R¹ stands for hydroxyl group. R³ together with R⁶ stands for anether group, R⁴ and R⁸ are mutually the same and stand for hydrogen andR⁵ and R⁷ are mutually the same and stand for methyl, or3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-12-O-methyl-azithromycin3,6-hemiketal of the general formula (I), wherein R¹ stands for hydroxylgroup, R³ together with R⁶ stands for an ether group, R⁴ and R⁷ aremutually the same and stand for hydrogen, and R⁵ and R⁸ are mutually thesame and stand for methyl.

Step 9

3-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-azithromycin3,6-hemiketal from the Step 8 is subsequently optionally subjected to areaction with ethylene carbonate in the presence of inorganic or organicbases, preferably potassium carbonate, in a reaction-inert solvent,preferably ethyl acetate, yielding3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-azithromycin3,6-hemiketal 11,12 cyclic carbonate of the general formula (I), whereinR¹ stands for hydroxyl group, R³ together with R⁶ stands for an ethergroup, R⁴ is hydrogen, R⁵ is methyl and R⁷ and R⁸ together with C-11 andC-12 carbon atoms stand for a cyclic carbonate.

Pharmaceutically acceptable addition salts, which are another object ofthe present invention, are obtained by a reaction of the novel compoundsof the general formula (I) with an at least equimolar amount of acorresponding inorganic or organic acid such as hydrochloric,hydroiodic, sulfuric, phosphoric, acetic, propionic, trifluoroacetic,maleic, citric, stearic, succinic, ethylsuccinic, methanesulfonic,benzene-sulfonic, p-toluenesulfonic, laurylsulfonic and similar acids,in a reaction-inert solvent. The addition salts are isolated byfiltration if they are insoluble in the reaction-inert solvent, byprecipitation with a nonsolvent or by evaporation of the solvent, mostfrequently by lyophilization.

Antibacterial in vitro activity of the novel compounds of the generalformula (I) and their pharmaceutically acceptable addition salts withinorganic or organic acids on a series of standard test-microorganismswas determined in a Mueller-Hinton medium (Difco-Laboratories, Detroit,Mich.) by a conventional method of double dilution in accordance withrecommendations of NCCLS (The National Committee for Clinical LaboratoryStandards). Each test microorganism was inoculated to the final inoculumsize of 5×10⁵ cfu/ml and the incubation was carried out in an anaerobicmanner at 37° C. during 18 hours. The MIC in the liquid medium wasdefined as the lowest concentration of an antibacterial agent inhibitingvisible growth in microdilutional containers. Control organisms wereobtained from ATCC (The American Type Culture Collection). All standardswere identified by a standard procedure and were storaged at −70° C. Theresults of 12-O-methyl-azithromycin on standard test microorganisms andclinical isolates in comparison with azithromycin are shown in Table 1and Table 2.

By determining the concentration of 12-O-methyl-azithromycin in serumafter a single oral dosis of 20 mg/kg on a group of 36 male rats in timeintervals from 0.25 to 24 hours it was established that the novelantibiotic was very fast absorbed in the serum. An analysis of the peakssuggested the existence of enterohepatic circulation. During 0.5 and 1hours a rapid drop of concentration took place, which was followed by arepeated increase. The maximum substance concentration was achievedafter 2 hours (Cmax 248.8 ng/ml). A secondary maximum was achieved 4hours after the application. The half-life was 5.2 hours and the totalAUC was 1993.4 h ng/ml.

TABLE 1 Antibacterial in vitro activity of 12-O-methyl-azithromycin onstandard strains in comparison with azithromycin MIC (mcg/ml)12-O-Methyl- Organism Azithromycin azithromycin Staphylococcus aureusATCC 6538 P 1 0.25 S. aureus ATCC 29213 0.25 0.25 S. eperidermidis ATCC12228 0.5 0.03 Micrococcus flavus ATCC 10240 0.5 0.12 M. luteus ATCC9341 0.06 0.03 Streptococcus faecalis ATCC 8043 0.5 0.25 Bacillussubtilis ATCC 6633 4 1 B. cereus ATCC 11778 1 0.25 Escherichia coli ATCC10536 1 0.5

TABLE 2 Antibacterial in vitro activity of 12-O-methyl-azithromycin on aseries of clinical isolates in comparison with azithromycin Organism MIC(μg/ml) (No. of strains) Compound Range 50% 90% Staph. aureus.Azithromycin 0.25-8 1 4 (77) 12-O-Methylazithromycin 0.12-2 0.25 1 S.epidermidis Azithromycin 0.25-16 0.25 8 (20) 12-O-Methylazithromycin0.12-8 0.25 4 Streptococcus Azithromycin 0.03-0.25 0.05 0.12 pneumoniae12-O-Methylazithromycin 0.03-0.12 0.03 0.12 (25) Enterococcus sp.Azithromycin 0.25-16 1 16 (35) 12-O-Methylazithromycin 0.12-8 0.5 8Haemophilus Azithromycin 0.12-0.5 0.25 0.5 influenzae12-O-Methylazithromycin 0.06-0.5 0.12 0.25 (40)

The process for the preparation of novel 3,6-hemiketals from the classof 9a-azalides is illustrated by the following examples, which in no waylimit the scope of the invention.

Preparation 1

2′-O,3′-N-Bis(bezyloxyearbonyl)-3′-N-demethyl-azithromycin A

To a solution of azithromycin (17 g, 0.0227 mole) in toluene (170 ml),NaHCO₃ (74.8 g, 0.890 mole) was added and then the reaction mixture washeated under stirring to reflux temperature (80-85° C.). To the reactionsuspension 102 ml of 50% benzyloxy-carbonyl chloride (104.04 g, 0.305mole) in toluene were added dropwise under stirring during 1 hour. Thereaction mixture was stirred at the same temperature for furher 2 hoursand left standing over night at room temperature. After filtration theprecipitate was rinsed with toluene (85 ml) and the toluene solution wasextracted twice with 0.25 N HCl (170 ml) and twice with 1.5% aqueousNaCl solution (170 ml). To toluene water was added (340 ml) (pH 3.1),the pH of the reaction mixture was adjusted with 6 N HCl to 2.0, thelayers were separated and the organic layer was further extracted threetimes with water (340 ml) under keeping the pH at 2.0. To combined waterextracts CH₂Cl₂ (125 ml) was added, the pH was adjusted with an aqueousNaOH solution (20%) to 10, the layers were separated and the aqueouslayer was again extracted with CH₂Cl₂(125 ml). The combined organicextracts were dried over K₂CO₃, filtered and evaporated at a reducedpressure, yielding 16.5 g of a thick oily residue, which was optionallypurified with low-pressure chromatography on a silica gel 60 column(230-400 mesh ASTM). For this purpose the crude product was dissolved inCH₂Cl₂ (20 ml) and applied to a silica gel column (50 g) under nitrogenpressure of 0.5 bar. In order to remove the residual benzylchloroformateand its disintegration products, CH₂Cl₂ (150 ml) was led through thecolumn and then by using the solvent system methylene chloride-methanol,9:1 (200 ml) and evaporating the fractions containingchromatographically homogeneous title product, there were obtained 11.53g of TLC pure 2′-O,3′-N-bis-(benzyloxycarbonyl)-N-demethyl-azitruomycinwith physical-chemical constants as described in U.S. Pat. No. 5,250,518of October 1993.

EXAMPLE 14″,11-O-Bis(trimethylsilyl)-2′-O,3′-N-bis(benzyloxycarbonyl)-3′-N-demethyl-azithromycin

To a solution of2′-O,3′-N-bis(benzyloxycarbonyl)-3′-N-demethyl-azithromycin (5.0 g,0.005 mole) in pyridine (50 ml), cooled to 0-5° C.,trimethylsilylimidazole (3.3 ml, 0.0226 mole) and trimethylsilylchloride(3.0 ml, 0.0179 mole) were added under nitrogen stream. The reactionmixture was stirred at the same temperature for 6 hours, n-hexane (60ml) and water (100 ml) were added, the layers were separated and theorganic layer was rinsed with a saturated NaHCO₃ solution (60 ml) andwater (60 ml). After drying over MgSO₄, filtration and evaporation ofthe solvent at a reduced pressure, 5.48 g of a white amorphousprecipitate were obtained, which were optionally purified bylow-pressure chromatography on a silica gel column using the systemCH₂Cl₂—CH₃OH, 9:1. The combining and evaporation of chromatographicallyhomogeneous fractions gave the title product with the followingphysical-chemnical constants:

TLC, Methylene chloride-methanol, 90:1 Rf 0.875 Ethylacetate-N-hexane-diethyl amine, 100:100:20 Rf 0.942

(IR (KBr) cm⁻¹: 3524, 2969, 2692, 1754, 1732, 1708, 1498, 1456, 1382,1335, 1252, 1168, 1116, 1060, 1005, 895, 841, 754, 696.

¹H NMR(300 MHz, CDCl₃) δ: 7.32-7.23 (Ph), 5.12, 4.98 (CH₂—Ph), 4.85(H-1″), 4.70 (H-1′), 4.65 (H-2′), 4.46 (H-3′), 4.26 (H-5″), 4.42 (H-3),3.72 (H-5′), 3.66 (H-11) 3.49, 3.47 (H-5), 3.20 (H-4″), 3.32, 3.18(3″-OCH₃), 2.83, 2.79 (3′-NCH₃), 2.7 (H-2), 2.64 (H-10), 2.35 (H-9a),2.33 (H-2″a), 2.11 (9a-NCH₃), 1.94 (H-9b), 1.91 (H-8), 1.64 (H-14a),1.94 (H-4), 1.50 (H-2″b), 1.50 (H-14b), 1.27, 1.25 (6-CH₃), 1.24(5″-CH₃), 1.19 (5′-CH₃), 1.12 (3″-CH₃), 1.16 (12-CH₃), 1.26 (2-CH₃),0.89 (10-CH₃), 0.95 (8-CH₃), 0.85 (14-CH₃), 1.02 (4-CH₃), 1.02 (4-CH₃),0.16 (11-OSi(CH₃)₃, and 0.13 /4″-OSi(CH₃)₃/.

¹³C NMR (75 MHz, CDCl₃) δ: 176.2 (C-1), 156.2, 156.4 (OCO), 154.5, 154.4(NCO), 136.7-127.5 (Ph), 100.2 (C-1′), 97.3 (C-1″), 83.9 (C-5), 80.7(C-4″), 75.0 (C-3), 75.0 (C-2′), 75.3 (C-6), 73.2 (C-3″), 69.4, 69.2,67.1, 66.8 (CH₂—Ph), 64.8 (C-5″), 62.3 (C-10), 54.8 (C-3′), 49.4, 49.2(3″-OCH₃), 46.2 (C-2), 38.5 (C-7), 39.4 (C-4), 34.2 (9a-NCH₃), 35.9,35.6 (C-2″), 36.2, 36.1 (C-4′), 29.0 (3′-NCH₃), 25.6 (C-8), 27.8(6-CH₃), 21.9 (3″-CH₃), 21.5 (8-CH₃), 20.7 (5′-CH₃), 23.4 (C-14), 18.4(5″-CH₃), 16.0 (2-CH₃), 11.6 (14-CH₃), 9.6, 9.5 (4-CH₃), 8.3 (10-CH₃),1.2 /11-OSi(CH₃)/₃ and 0.67/4″-OSi(CH₃)₃/.

ES-MS 1147

EXAMPLE 2 3′-N-Demethyl-12-O-methyl-azithromycin

To a solution of the product from Example 1 (1.0 g, 0.0009 mole) inN,N-dimethyl-formamide (20 ml) methyl iodide (0.43 ml, 0.0069 mole) and60% sodium hydride (0.23 g, 0.0058 mole) were gradually added during 3hours at room temperature. The reaction mixture was stirred for further30 minutes at the same temperature, the reaction was stopped by theaddition of triethyl amine (2 ml), it was transferred into a mixture of10% aqueous NaHCO₃ solution (50 ml) and water (50 ml) and extracted withethyl acetate. The combined organic extracts were rinsed with asaturated NaCl solution and water, dried over MgSO₄, filtered andevaporated at a reduced pressure, yielding 0.93 g of a yellowprecipitate [Rf 0.832, methylene chloride-methanol, 90:1; IR(KBr)cm⁻¹:3516, 1752, 1732, 1705, 1456, 1382, 1336, 1253, 1169, 1116, 1062, 1004,896, 840, 754, 696]. The product was dissolved in ethanol (20 ml),NaOAc/HOAc buffer with pH 5 (0.17 ml acetic acid, 0.263 g sodiumacetate, 0.22 ml ethanol and 1 ml water) and Pd/C 10% (0.6 g) wereadded, and the reaction mixture was hydrogenated under stirring for 5hours in an autoclave at a hydrogen pressure of 5 bars, The catalyst wasfiltered off, the filtrate was evaporated to a thick syrup, CH₂Cl₂ (10ml) and water (15 ml) were added, the pH of the mixture was adjustedwith 2 N HCl to 4, the layers were separated and the aqueous layer was,upon adjustment to pH 9.5 with 20% NaOH, extracted with CH₂Cl₂ (3×10ml). The combined organic extracts were dried over K₂CO₃, filtered andevaporated. The precipitate was dissolved in isopropanol (10 ml), water(10 ml) and some drops of formic acid were added and it was stirred for30 minutes at room temperature, extracted with isopropyl acetate at pH9.5, which upon evaporation at a reduced pressure yielded 0.43 g of thetitle product with the following physical-chemical constants:

IR (KBr) cm⁻¹: 3672, 3496, 2962, 1727, 1458, 1375, 1343, 1280, 1263,1118, 1085, 1048, 1005, 998.

¹³C NMR (75 MHz, CDCl₃) δ: 177.4 (C-1), 102.7 (C-1′), 95.5 (C-1″), 83.4(C-5), 79.7 (C-12), 78.0 (C-3), 76.6 (C-11), 74.0 (C-13), 73.9 (C-6),74.3 (C-2′), 73.0 (C-3″), 68.8 (C-9), 65.7 (C-5″), 60.1 (C-3′), 61.2(C-10), 52.8 (12-OCH₃), 49.8 (3″-OCH₃), 45.5 (C-2), 41.5 (C-4), 33.1,3′-NCH₃, 36.8 (9a-NCH₃), 35.1 (C-2″), 28.8 (C-4′), 27.0 (C-8).

EI-MS m/z 748.

Example 3 12-O-Methyl-azithromycin

To a solution of 3′-N-demethyl-12-O-methyl-azithromycin from Example 2(0.43 g, 0.0006 mole) in CHCl₃ (20 ml), formaldehyde (37%) (0.047 ml,0.0006 mole) and formic acid (98-100%) (0.042 ml, 0.0011 mole) wereadded. The reaction mixture was stirred for 3 hours under reflux, cooledto room temperature, poured onto water (20 ml) and upon adjustment of pHto 4.0, the layers were separated and the aqueous layer was extractedtwo more times with CHCl₃. To the aqueous layer CHCl₃ was added, the pHwas adjusted to 9.5 (2N NaOH), the layers were separated and the aqueousone was extracted two more times with CHCl₃. The combined organicextracts at pH 9.5 were dried (K₂CO₃) and evaporated, yielding 0.38 g ofthe title product, which was, if necessary, purified by a chromatographyon a silica gel column using the system CH₂Cl₂—CH₃OH-conc.NH₄OH, 90:9:1.

TLC, Methylene chloride-methanol-conc. ammonia, 90:9:0.5 Rf 0.363 Ethylacetate-N-hexane-diethyl amine, 100:100:20 Rf 0.745

IR (KBr) cm⁻¹: 3499, 2972, 2940, 1736, 1633, 1460, 1381, 1259, 1168,1110, 1059, 1082, 1054, 1013, 999.

¹H NMR (300 MHz, CDCl₃) δ: 5.39 (H-13), 5.00 (H-1″), 4.43 (H-1′), 4.32(H-3), 4.06 (H-5″), 3.68 (H-11), 3.65 (H-5), 3.51 (H-5′), 3.38(12-OCH₃), 3.32 (3″-OCH₃), 3.24 (H-2′), 3.02 (H-4″), 2.73 (H-2), 2.69(H-10), 2.49 (H-3′), 2.34 (H-2″a), 2.31 (H-9a), 2.29 /3′N(CH₃)₂/, 2.30(9a-NCH₃), 2.12 (H-9b), 2.04 (H-4), 2.01 (H-8), 1.73 (H-14a), 1,68(H-4′a), 1.66 (H-7a), 1.56 (H-2″b), 1.52 (H-14b), 1.36 (H-7b), 1.29(6-CH₃), 1.21 (2-CH₃), 1.30 (5″-CH₃), 1.24 (H-4′b), 1.23 (3″-CH₃), 1.22(5′-CH₃), 1.09 (12-CH₃), 1.29 (4-CH₃), 1.09 (10-CH₃), 0.92 (8-CH₃), 0.93(14-CH3).

¹³C NMR (75 MHz, CDCl₃) δ177.5 (C-1), 103.1 (C-1′), 95.2 (C-1″), 83.6(C-5), 79.2 (C-12), 78.1 (C-3), 76.6 (C-11), 74.7 (C-13), 73.8 (C-6),70.9 (C-2′), 68.8 (C-9), 65.6 (C-5″), 65.7 (C-3′), 61.6 (C-10), 52.8(12-OCH₃), 49.4 (3″-OCH₃), 45.1 (C-2), 43.0 (C-7), 41.8 (C-4), 40.4/3′N(CH₃)₂/, 36.8 (9a-NCH₃), 35.0 (C-2″), 29.0 (C-4′), 26.9 (C-8), 26.9(6-CH₃), 22.0 (8-CH₃), 22.0 (C-14), 21.6 (3″-CH₃), 21.3 (5′-CH₃), 18.1(5″-CH₃), 16.9 (12-CH₃), 14.6 (2-CH₃), 11.0 (14-CH₃), 9.6 (4-CH₃), 9.4(10-CH₃).

EXAMPLE 43-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-12-O-methyl-azithromycin

In 0.25 N hydrochloric acid (80 ml) 12-O-methyl-azithromycin (1.7 g,0.0022 mole) from Example 3 was dissolved and it was left standing for24 hours at room temperature. To the reaction mixture CH₂Cl₂ (pH 1.8)was added, the layers were separated and the aqueous one was extractedtwo more times with CH₂Cl₂. To the aqueous layer again CH₂Cl₂ was added,the pH of the rmixture was adjusted with conc. NH₄OH to 9.0, the layerswere separated and the aqueous one was extracted with CH₂Cl₂. Thecombined organic extracts at pH 9.0 were rinsed with 10% aqueous NaHCO₃solution and water, dried over K₂CO₃ and evaporated, yielding 1.25 g ofthe title product with the following physical-chemical constants:

TLC, Methylene chloride-methanol-conc. ammonia, 90:9:0.5 Rf 0.315 Ethylacetate-N-hexane-diethyl amine, 100:100:20 Rf 0.594

IR (KBr) cm⁻¹: 3450, 2971, 2933, 1711, 1648, 1460, 1381, 1272, 1261,1171, 1113, 1078, 1049.

¹H NMR (300 MHz, CDCl₃) δ: 5.32 (H-13), 4.47 (H-1′), 3.78 (H-3), 3.66(H-11), 3.58 (H-5), 3.58 (H-5′), 3.41 (12-OCH₃), 3.28 (H-2′), 2.67(H-2), 2.80 (H-10), 2.35 (H-3′), 2.53 (H-9a), 2.27 /3′N(CH₃)₂/, 2.37(9a-NCH₃), 2.07 (H-9b), 2,27 (H-4), 1.92 (H-8), 1.74 (H-14a), 1.68(H-4′a), 1.59 (H-7a), 1.63 (H-14b), 1.51 (H-7b), 1.31 (6CH₃), 1.31(2-CH₃), 1.29 (H-4′b), 1.26 (5′-CH₃), 1.08 (12-CH₃), 1.05 (4-CH₃), 1.19(10-CH₃), 0.93 (8-CH₃), 0.92 (14-CH₃).

¹³C NMR (75 MHz, CDCl₃) δ: 177.2 (C-1), 106.4 (C-¹′), 94.7 (C-5), 78.0(C-12), 79.0 (C-3), 78.3 (C-11), 75.1 (C-13), 72.9 (C-6), 70.2 (C-2′),70.3 (C-9), 65.3 (C-3′), 62.1 (C-10), 52.5 (12-OCH₃), 44.3 (C-2), 41.8(C-7), 35.7 (C-4), 39.9 /3′N(CH₃)₂/, 36.5 (9a-NCH₃), 27.9 (C-4′), 26.4(C-8), 25.5 (6-CH₃), 20.8 (8-CH₃), 20.7 (C-14), 20.8 (5′-CH₃, 16.1(12-CH₃), 15.7 (2-CH₃), 10.3 (14-CH₃), 7.6 (4-CH₃), 7.2 (10-CH₃).

EXAMPLE 53-De(2,6-dideoxy-3-C-methyl-3-O-methyl-δ-L-ribohexopyranosyl-oxy)-3-oxy-12-O-methyl-azithromycin-2′-O-acetate

To a solution of3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-12-O-methyl-azithromycin(1.3 g, 0.0022 mole) from Example 4 in CH₂Cl₂ (20 ml), NaHCO₃ (0.754 g,0.009 mole) and acetic acid anhydride (0.221 ml, 0.0023 mole) were addedand then it was stirred for 10 hours at room temperature. After standingover night saturated NaHCO₃ solution was added to the reaction mixture,the layers were separated and the aqueous one was extracted with CH₂Cl₂.The combined organic extracts were rinsed with a saturated NaHCO₃solution and water, dried over K₂CO₃, filtered and evaporated, yielding1.29 g of a white amorphous precipitate.

TLC, Methylene chloride-methanol-conc. ammonia, 90:9:0.5 Rf 0.489 Ethylacetate-N-hexane-diethyl amine, 100:100:20 Rf 0.661

IR (KBr) cm⁻¹: 3448, 2974, 1749, 1718, 1637, 1458, 1377, 1242, 1169,1115, 1045.

¹H NMR (300 MHz, CDCl₃) δ: 5.23 (H-13), 4.72 (H-2′), 4.70 (H-1′), 3.59(H-11), 3.56 (H-5), 3.52 (H-3), 3.43 (H-5′), 3.33 (12-OCH₃), 2.72(H-10), 2.71 (H-3′), 2.61 (H-2), 2.42 (H-9a), 2.30 (9a-NCH₃), 2.20/3′N(CH₃)₂/, 2.12 (H-4, 1.99 (2′-COCH₃), 1.96 (H-9b), 1.80 (H-8), 1.67(H-14a), 1.67 (H-4′a), 1.58 (H-14b), 1.47 (H-7a), 1.31 (H-4′b), 1.21(2-CH₃), 1.18 (H-7b), 1.16 (5′-CH₃), 1.15 (6-CH₃), 1.10 (10-CH₃), 0.97(12-CH₃), 0.86 (14-CH₃), 0.84 (8-CH₃), 0.81 (4-CH₃).

¹³C NMR (75 MHz, CDCl₃) δ: 176.5 (C-1), 169.4 (2′-COCH₃), 98.6 (C-1′),84.3 (C-5), 77.3 (C-12), 78.3 (C-3), 76.7 (C-11), 74.6 (C-13), 72.4(C-6), 70.7 (C-2′), 69.9 (C-9), 62.2 (C-3′), 62.3 (C-10), 51.9(12-OCH₃), 43.0 (C-2), 40.1 (C-7), 35.2 (C-4), 39.6 /3′N(CH₃)₂/, 35.9(9a-NCH₃), 30.0 (C-4′), 25.4 (C-8), 25.2 (6-CH₃), 20.6 (2′COCH₃), 20.4(8-CH₃), 20.0 (C-14), 20.2 (5′-CH₃), 15.9 (12-CH₃), 15.2 (2-CH₃), 9.7(14-CH₃), 7.0 (4-CH₃), 6.4 (10-CH₃).

EXAMPLE 63-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexyopyranosyl-oxy)-12-O-methyl-azithromycin3,6-hemiketal-2 ′-O-acetate

To a solution of3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-δ-L-ribohexopyranosyl-oxy)-3-oxy-12-O-methyl-azithromycin2′-O-acetate (1.3 g, 0.0020 mole) from Example 5 in CH₂Cl₂ (15 ml),dimethyl sulfoxide (4.35 ml) andN,N-dimethyl-amino-propyl-ethyl-carbodiimide (4.55 g) were added. Thereaction mixture was cooled to 15° C. and then under stirring andkeeping the temperature at 15° C. a solution of pyridiriumtrifluoroacetate (4.61 g, 0.0234 mole) in CH₂Cl₂ (10 ml) was addeddropwise over 30 minutes. The temperature of the reaction mixture wasgradually increased to room temperature and it was stirred for firther 2hours, whereupon the reaction was stopped by the addition of a saturatedNaCl solution (25 ml). After alkalizing with 2 N NaOH to 9.5, thereaction mixture was extracted with CH₂Cl₂, the organic extracts wererinsed with a saturated NaCl solution, NaHCO₃ and water and dried overK₂CO₃. The evaporation of CH₂Cl₂ at a reduced pressure gave 1.78 g of anoily residue.

TLC, Methylenechloride-methanol-conc. ammonia, 90:9:0.5 Rf 0.176 Ethylacetate-N-hexane-diethyl amine, 100:100:20 Rf 0.861

EXAMPLE 73-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-12-O-methyl-azithromycin3,6-hemiketal

A solution of3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-(α-L-ribohexopyranosyl-oxy)-12-O-methyl-azithromycin2′-O-acetate (1.78 g) from Example 6 in methanol (50 ml) was leftstanding for 24 hours at room temperature. Methanol was evaporated at areduced pressure, the obtained residue (1.65 g) was purified bylow-pressure chromatography on a silica gel column using the systemmethylene chloride-methanol-conc. ammonia, 90:9:0.5. By evaporating thecombined extracts with Rf 0.082 there was obtained chromatographicallyhomogeneous3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-δ-L-ribohexopyranosyl-oxy)-12-O-methyl-azithromycin-3,6-herniketalwith the following physical-chemical constants:

TLC, Methylene chloride-methanol-conc. ammonia, 90:9:0.5 Rf 0.082 Ethylacetate-N-hexane-diethyl amine, 100:100:20 Rf 0.624

IR (CDCl₃) cm⁻¹: 3450, 2956, 2940, 1718, 1678, 1631, 1459, 1383, 1278,1198, 1117, 1068, 1048, 1014, 963.

¹H NMR (300 MHz, CDCl₃) δ: 5.49 (H-13), 4.21 (H-1′), 3.83 (H-11), 3.75(H-5),3.52 (H-5′), 3.43 (12-OCH₃), 3.25 (H-2′), 2.59 (H-2), 2.93 (H-10),2.50 (H-3′), 2.61 (H-9a), 2.29 /3′N(CH₃)₂)/, 2.40 (9a-NCH₃), 2.10(H-9b), 2.06 (H-4), 1.88 (H-8), 1.77 (H-14a), 1.67 (H-4′a), 1.61 (H-7a),1.64 (H-14b), 1.33 (H-7b), 1.31 (6-CH₃), 1.05 (2-CH₃), 1.27 (H-4′b),1.26 (5′-CH₃), 1.08 (12-CH₃), 1.05, (4-CH₃), 1.19 (10-CH₃), 0.92(8-CH₃), 0.93 (14-CH₃).

¹³C NMR (75 MHz, CDCl₃) δ: 176.2 (C-1), 105.8 (C-1′), 94.6 (C-5), 78.3(C-12), 102.7 (C-3), 71.2 (C-11), 74.8 (C-13), 82.9 (C-6), 69.6 (C-2′),64.5 (C-9), 65.1 (C-3′), 60.7 (C-10), 52.2 (12-OCH₃), 49.2 (C-2), 41.4(C-7), 48.6 (C-4), 40.0 /3′N(CH₃)₂/, 40.5 (9a-NCH₃), 28.2 (C-4′), 29.1(C-8), 26.5 (6-CH₃), 21.5 (8-CH₃), 21.6 (C-14), 20.8 (5′-CH₃), 16.3(12-CH₃), 13.6 (2-CH₃), 10.7 (14-CH₃), 12.8 (4-CH₃), 10.7 (10-CH₃).

EXAMPLE 84″-O-Trimethylsilyl-2′-O-3′-N-bis(benzyloxycarbonyl)-3′-N-demethyl-azithromycin

To a solution of2′-O,3′-N-bis(benzyloxycarbonyl)-3′-N-demethyl-azithromycin (5 g, 0.005mole) in pyridine (30 ml) cooled to 0-5° C., trimethylsilyl imidazole(1.46 ml, 0.01 mole) and trimethylsilyl chloride (1.64 ml, 0.01 mole)were added under a nitrogen stream. The reaction mixture was stirred for1 hour at the same temperature, n-hexane (50 ml) and water (25 ml) wereadded, the layers were separated and the organic one was rinsed with asaturated NaHCO₃ solution (25 ml) and water (25 ml). After drying overMgSO₄, filtration and evaporation of the solvent at a reduced pressurethere was obtained an amorphous precipitate (3.65 g), which wasoptionally purified by low-pressure chromatography on a silica gelcolumn using the system methylene chloride-methanol-conc. ammonia,90:9:0.5. By combining and evaporating the chromatographicallyhomogeneous fractions with Rf 0.670 there was obtained the title productwith the following physical-chemical constants:

TLC, Methylene chloride-methanol, 90:1 Rf 0.525 Ethylacetate-N-hexane-diethyl amine, 100:100:20 Rf 0.862

IR (KBr) cm⁻¹: 3502, 2969, 2938, 1753, 1732, 1708, 1454, 1383, 1365,1254, 1169, 1118, 1063, 1001, 897, 839, 754, 696.

¹H NMR (300 MHz, CDCl₃) δ: 7.34-7.26 (Ph), 5.13, 5.09, (CH₂—Ph), 5.07(H-1″), 4.78 (H-1′), 4.68 (H-13), 4.66 (H-2′), 4.55 (H-3′), 4.22 (H-5″),4.13 (H-3), 3.96 (H-5′), 3.65 (H-11), 3.58, 3.54 (H-5), 3.15 (H-4″),3.37, 2.99 (3″-OCH₃), 2.85, 2.81 (3′-NCH₃), 2.70 (H-2), 2.68 (H-10),2.54 (H-9a), 2.35 (H-2″a), 2.31 (9a-NCH₃), 2.04 (H-9b), 1.97 (H-8), 1.90(H-14a), 1.85 (H-4), 1.62 (H-7a), 1.50 (H-2″b), 1.44 (H-14b), 1.28, 1.27(6-CH₃), 1.23 (5″-CH₃), 1.16 (5′-CH₃), 1.15 (H-7b), 1.04 (3″-CH₃), 1.15(12-CH₃), 1.10 (2-CH₃), 1.10 (10-CH₃), 0.92 (8-CH₃), 0.89 (14-CH₃), 1.10(4-CH₃).

¹³C NMR (75 MHz, CDCl₃) δ: 178.8 (C-1), 156.6, 156,3 (OCO), 154.7, 154.6(NCO), 136.8-127-5 (Ph), 99.2 (C-1′), 94.8 (C-1′″), 83.2, 83.1 (C-5),80.5, 80.4 (C-4″), 77.3 (C-3), 75.1, 75.0 (C-2′), 74.1 (C-12), 73.8(C-11), 73.2 (C-6), 73.2 (C-3 ″), 69.2, 69.0 67.2, 66.8 (CH₂-Ph), 64.8(C-5″), 62.2 (C-10), 54.6 (C-3′), 49.3, 48.8 (3″-OCH₃), 44.7 (C-2), 41.5(C-7), 41.1 (C-4), 36.1 (9a-NCH₃), 35.1, 35.0 (C-2″), 36.3, 35.7 (C-4′),28.4 (3′-NCH₃), 26.3 (C-8), 26.8 (6-CH₃), 22.1 (3″-CH₃), 21.6 (8-CH₃),21.4 (5′-CH₃), 21.0 (C-14), 18.7 (5″-CH₃), 15.9 (2-CH₃), 14.5 (12-CH₃),11.0 (14-CH₃), 8.5 (4-CH₃), 7.1 (10-CH₃), 0.63/4″-OSi(CH₃)₃/.

ES-MS 1075.

EXAMPLE 9 11-O-methyl-azithromycin and 12-O-methyl azithromycin

To a solution of the product from Example 8 (3.0 g, 0.0028 mole) inN,N-dimethylformamide (50 ml), methyl iodide (1.29 ml, 0.0207 mole) and60% sodium hydride (0.69 g, 0.0174 mole) were gradually added over 3hours at room temperature. The reaction mixture was stirred for 1 hourat the same temperature, the reaction was stopped by addition oftriethylamine (5 ml), it was transferred into a mixture of 10% aqueousNaHCO₃ solution (100 ml) and water (100 ml) and extracted with ethylacetate. The combined organic extracts were rinsed with a saturated NaClsolution and water and dried over MgSO₄, filtered and evaporated at areduced pressure yielding 2.9 g of a mixture of products, which wasoptionally purified by low-pressure chromatography on a silica gelcolumn using the system methylene chloride-methanol, 90:1, yielding achromatographically homogeneous4″-O-trimethylsilyl-2′-O-3′-N-bis(benzyloxy-carbonyl)-3′-N-demethyl-11-O-methyl-azithromycinwith Rf 0.745 [IR (KBr): 3452, 2969, 1752, 1736, 1706, 1455, 1382, 1332,1254, 1169, 1117, 1063, 1002, 914, 897, 840, 754, 697] and4″-O-trimethylsilyl-2′-O-3′-N-bis(benzyloxy-carbonyl)-3′-N-demethyl-12-O-methyl-azithroinycinwith Rf 0.485 [IR (KBr): 3450, 2958, 1754, 1718, 1708, 1458, 1383, 1252,1168, 1068, 1010, 896, 842, 753, 695].

The obtained mixture was dissolved in ethanol (50 ml), NaOAc/HOAc bufferwith pH 5 (0.51 ml HOAc, 0.789 g NaOAc, 0.66 ml ethanol and 3 ml water)and 10% Pd/C (1.5 g) were added and the mixture was hydrogenated understining for 8 hours in an autoclave at a hydrogen pressure of 5 bars.The catalyst was filtered off, the filtrate was evaporated to a thicksyrup, water (50 ml) and CHCl₃ (50 ml) were added and the product wasisolated by a pH gradient extraction at pH 4.0 and 9.5. The combinedorganic extracts at pH 9.5 were dried over K₂CO₃ and evaporated to anamorphous precipitate. The precipitate was dissolved in isopropanol (20ml), water (20 ml) and some drops of formic acid were added and it wasstirred for 30 minutes at room temperature, extracted with isopropylacetate at pH 9.5, dried over sodium sulfate and evaporated at a reducedpressure. The obtained product was dissolved in CHCl₃ (50 ml),formaldehyde (37%) (0.24 ml) and formic acid (98-100%) (0.22 ml) wereadded. The reaction mixture was stirred for 3 hours under reflux, cooledto room temperature, poured onto water (20 ml) and after adjusting thepH to 4.0 the layers were separated and the aqueous one was extractedtwo more times with CHCl₃. To the water layer CHCl₃ was added, pH wasadjusted to 9.5 (2 N NaOH), the layers were separated and the aqueousone was extracted two more times with CHCl₃. The combined organicextracts at pH 9.5 were dried (K₂CO₃) and evaporated, yielding 1.25 g ofa precipitate, which was chromatographed on a silica gel column usingthe system methylene chloride-methanol-conc. ammonia, 90:9:1, yielding0.40 g of chromatographically homogeneous 11-O-methyl-azithromycin withphysical-chemical constants as given in U.S. Pat. No. 5,250,518 ofOctober 1993 and 0.52 g of 12-O-methyl-azithromycin withphysical-chemical constants as given in Example 3.

EXAMPLE 103-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-11-O-methyl-azithromycin

In methanol (30 ml) 11-O-methyl-azithromycin (1.5 g) was dissolved, 0.25N hydrochloric acid (50 ml) was added and it was left standing for 24hours at room temperature. Methanol was evaporated, to the reactionmixture CDCl₃ (pH 1.9) was added, the layers were separated and theaqueous one was extracted two more times with CDCl₃. The aqueoussolution was alkalized to pH 9.5 and extracted with CDCl₃ The combinedorganic exctracts at pH 9.5 were dried over K₂CO₃ and evaporated,yielding 0.95 g of the title product, which was optionally purified bylow-pressure chromatography on a silica gel column using the solventsystem methylene chloride-methanol-conc. ammonia, 90:9:0.5, yielding achromatographically homogeneous title product with the followingphysical-chemical constants:

TLC, Methylene chloride-methanol-conc. ammonia, 90:9:0.5 Rf 0.382 Ethylacetate-N-hexane-diethyl amine, 100:100:20 Rf 0.594

IR (KBr) cm⁻¹: 3448, 2972, 2937, 1730, 1638, 1458, 1377, 1165, 1113,1078, 1050.

¹H NMR (300 MHz, CDCl₃) δ: 4.97 (H-13), 4.52 (H-1′), 3.76 (H-3), 3.70(11-OCH₃), 3.59 (H-5), 3.54 (H-5′), 3.42 (H-11), 3.29 (H-2′), 2.68(H-2), 2.70 (H-10), 2.58 (H-3′), 2.46 (H-9a), 2.35 (H-4), 2.29/3′N(CH₃)₂/, 2.30 (9a-NCH₃), 2.11 (H-9b), 1.94 (H-14a), 1.89 (H-8), 1.70(H-4′a), 1.66 (H-7a), 1.54 (H-7b), 1.52 (H-14b), 1.33 (6-CH₃), 1.30(2-CH₃), 1.27 (H-4′b), 1.25 (5′-CH₃), 1.12 (12-CH₃), 1.10 (4-CH₃), 1.06(10-CH₃), 0.92 (8-CH₃), 0.86 (14-CH₃).

¹³C NMR (75 MHz, CDCl₃) δ175.7 (C-1), 106.1 (C-1′), 94.7 (C-5), 74.2(C-12), 78.1 (C-3), 86.0 (C-11), 77.1 (C-13), 72.8 (C-6), 70.2 (C-2′),70.9 (C-9), 65.4 (C-3′), 62.9 (C-10), 62.0 (11-OCH₃), 44.1 (C-2), 42.5(C-7), 35.3 (C-4), 39.9 /3′N(CH₃)₂/, 36.2 (9a-NCH₃), 28.0 (C-4′), 26.7(C-8), 25.8 (6-CH₃), 20.9 (8-CH₃), 21.2 (C-14), 20.8 (5′-CH₃), 16.8(12-CH₃), 15.6 (2-CH₃), 10.3 (14-CH₃), 7.7 (4-HC₃), 6.8 (10-CH₃).

EXAMPLE 113-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-11-O-methyl-azithromycin2′-O-acetate

To a solution of3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-11-O-methyl-azithromycin(0.89 g) from Example 10 in CH₂Cl₂ (25 ml), NaHCO₃ (0.52 g) and aceticacid anhydride (0.15 ml) were added, the reaction mixture was stirredfor 10 hours at room temperature, left standing over night and thenisolated by means of extraction with CH₂Cl₂ as described in Example 5,yielding 0.65 g of a white amorphous precipitate.

TLC, Methylene chloride-methanol-conc. ammonia, 90:9:0.5 Rf 0.426 Ethylacetate-N-hexane-diethyl amine, 100:100:20 Rf 0.670

IR (KBr) cm⁻¹ 3525, 3475, 2968, 2937, 1724, 1647, 1458, 1376, 1265,1168, 1113, 1081, 1050.

EXAMPLE 123-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-11-O-methyl-azithromycin3,6-hemiketal 2′-O-acetate

To a solution of3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-11-O-methyl-azithromycin2′-O-acetate (0.65 g) from Example 11 in CH₂Cl₂ (20 ml), dimethylsulfoxide (0.94 ml) and N,N-dimethyl-aminopropyl-ethyl-carbodiimide(1.16 g) were added. The reaction mixture was cooled to 15° C. and then,under stirring and maintaining the temperature at 15° C., a solution ofpyridinium trifluoroacetate (1.15 g) in CH₂Cl₂ (5 ml) was graduallyadded dropwise over 30 minutes. The temperature of the reaction mixturewas raised to room temperature, it was stirred for further 4 hours andthen a product was isolated according to the process described inExample 6, yielding 0.6 g of the title product.

TLC, Methylene chloride-methanol-conc. ammonia, 90:9:0.5 Rf 0.606 Ethylacetate-N-hexane-diethyl amine, 100:100:20 Rf 0.861

EXAMPLE 133-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-11-O-methyl-azithromycin3,6-hemiketal

A solution of3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-11-O-methyl-azithromycin3,6-hemiketal 2′-O-acetate (0.6 g) from Example 12 in methanol (40 ml)was left standing for 24 hours at room temperature. Methanol wasevaporated at a reduced pressure, the obtained residue (0.53 g) waspurified by low-pressure chromatography on a silica gel column using thesystem methylene chloride-methanol-conc. ammonia, 90:9:1.5. Byevaporation of the combined extracts with Rf 0.670 there were obtained0.22 g of chromatographically homogeneous3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-11-O-methyl-azitromycin3,6-heminketal with the following physical-chemical constants:

IR (CDCl₃) cm⁻¹: 3471, 2975, 1715, 1638, 1458, 1382, 1196, 1117, 1049,1013, 963.

¹H NMR (300 MHz, CDCl₃) δ: 5.01 (H-13), 4.22 (H-1′), 3.80 (H-5), 3.50(H-5′), 3.45 (11-OCH₃), 3.25 (H-2′), 2.63 (H-2), 2.49 (H-3′), 2.77(H-9a), 2.29 /3′N(CH₃)₂/, 2.20 (9a-NCH₃), 2.24 (H-9b), 2.09 (H-4), 1.85(H-8), 1.83 (H-14a), 1.66 (H-4′a), 1.73 (H-14b), 1.36 (6-CH₃), 1.31(2-CH₃), 1.26 (H-4′b), 1.21 (5′-CH₃), 1.25 (4-CH₃), 1.01 (10-CH₃), 1.03(8-CH₃), 0.81 (14-CH₃).

¹³C NMR (75 MHz, CDCl₃) δ: 177.0 (C-1), 106.2 (C-1′), 102.1 (C-3), 93.9(C-5), 86.1 (C-11), 81.9 (C-6), 69.7 (C-2′), 64.9 (C-9), 65.8 (C-3′),62.1 (C-10), 61.9 (11-OCH₃), 49.6 (C-2), 43.3 (C-7), 40.1 /3′N(CH₃)₂/,28.1 (C-4′), 28.7 (C-8), 25.5 (6-CH₃), 20.9 (5′-CH₃), 14.0 (2-CH₃), 11.7(14-CH₃), 12.3 (4-CH₃), 8.5 (10-CH₃).

EXAMPLE 143-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-azithromycin

3-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-azithromycinwas prepared from azithromycin according to the process by Djokićet al.from U.S. Pat. No. 4,886,792 of December 1989, Example 3. By separationon a silica gel column using the solvent system methylenechloride-methanol-conc. ammonia, 90:9:0.5, there was obtained achromatographically homogeneous product with the followingphysical-chemical constants:

TLC, Ethyl acetate-triethyl amine, 95:5 Rf 0.371

IR (KBr) cm⁻¹: 3438, 2973, 2938, 1713, 1655, 1459, 1378, 1350, 1260,1172, 1113, 1078, 1044, 957.

¹H NMR (300 MHz, CDCl₃) δ: 4.72 (H-13), 4.47 (H-1′), 3.78 (H-3), 3.58(H-5), 3.56 (H-5′), 3.65 (H-11), 3.27 (H-2′), 2.66 (H-2), 2.74 (H-10),2.52 (H-3′), 2.49 (H-9a), 2.28 (H-4), 2.26 /3′N(CH₃)₂/, 2.37 (9a-NCH₃),2.06 (H-9b), 1.90 (H-14a), 1.90 (H-8), 1.67 (H-4′a), 1.62 (H-7a), 1.47(H-7b), 1.53 (H-14b), 1.32 (6-CH₃), 1.30 (2-CH₃), 1.28 (H-4′b), 1.26(5′-CH₃), 1.07 (12-CH₃), 1.06 (4-CH₃), 1.12 (10-CH₃), 0.92 (8-CH₃), 0.88(14-CH₃).

¹³C NMR (75 MHz, CDCl₃) δ: 178.8 (C-1), 106.6 (C-1′), 94.7 (C-5), 72.9(C-12), 79,2 (C-3), 75.5 (C-11), 77.1 (C-13), 74.0 (C-6), 70.3 (C-2′),70.6 (C-9), 65.4 (C-3′), 62.2 (C-10), 44.2 (C-2), 41.7 (C-7), 35.6(C-4), 39.9 /3′N(CH₃)₂/, 36.8 (9a-NCH₃), 27.7 (C-4′), 26.3 (C-8), 25.5(6-CH₃), 20.8 (8-CH₃), 20.5 (C-14), 20.9 (5′-CH₃), 15.7 (12-CH₃), 15.8(2-CH₃), 10.5 (14-CH₃), 7.5 (4-CH₃), 7.3 (10 -CH₃).

EXAMPLE 153-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-azithromycin2′-O-acetate

To a solution of3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-3-oxy-azithromycin(10 g) from Example 14 in CH₂Cl₂ (150 ml), NaHCO₃ (5.84 g) and aceticacid anhydride (1.68 ml) were added. The reaction mixture was stirredfor 12 hours at room temperature, left standing over night and thenisolated according to the process described in Example 5, yielding 11.21g of an amorphous precipitate with the following physical-chemicalconstants:

TLC Ethyl acetate-triethyl amine, 95:5 Rf 0.547

IR (KBr) cm⁻¹: 3485, 2973, 2937, 1748, 1716, 1648, 1459, 1376, 1240,1170, 1114, 1081, 1045, 956.

¹H NMR (300 MHz, CDCl₃) δ: 4.71 (H-13), 4.79 (H-2′), 4.71 (H-1′), 3.84(H-3), 3.61 (H-5), 3.50 (H-5′), 3.68 (H-11), 2.73 (H-10), 2.70 (H-2),2.70 (H-3′), 2.48 (H-9a), 2.27 (H-4), 2.26 /3′N(CH₃)₂/ , 2.36 (9a-NCH₃),2.07 (COCH₃), 2.05 (H-9b), 1.90 (H-14a), 1.90 (H-8), 1.78 (H-4′a), 1.56(H-7a), 1.24 (H-7b), 1.54 (H-14b), 1.23 (6-CH₃), 1.29 (2-CH₃), 1.32(H-4′b), 1.24 (5′-CH₃), 1.11 (10-CH₃), 1.06 (12-CH₃), 0.90 (4-CH₃), 0.89(8-CH₃), 0.88 (14-CH₃).

EXAMPLE 163-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-azithromycin-3,6-hemiketal2′-O-acetate

To a solution of3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribo-hexopyranosyl-oxy)-3-oxy-azithromycin2′-O-acetate (5.6 g) from Example 15 in CH₂Cl₂ (100 ml), dimethylsulfoxide (12.34 ml) and N,N-dimethyl-aminopropyl-ethyl-carbodiimide(15.05 g) were added. The reaction mixture was cooled to 15° C. andthen, under stirring and maintaining the temperature at 15° C., asolution of pyridinium trifluoracetate (15.04 g) in CH₂Cl₂ (30 ml) wasadded gradually drop by drop over 30 minutes. The temperature of thereaction mixtue was raised to room temperature, it was kept stirring forfurther 4 hours and then a product was isolated in accordance with theprocess described in Example 6, yielding 5.26 g of the title product.

TLC Ethylacetate-trietyl amine, 95:5 Rf 0.675

EXAMPLE 173-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-azithromycin3,6-hemiketal

A solution of3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-azithromycin-3,6-hemiketal2′-O-acetate (5.2 g) from Example 16 in methanol (100 ml) was leftstanding for 16 hours at room temperature. Methanol was evaporated at areduced pressure and the obtained product was purified by low-pressurechromatography on a silica gel column using the system methylenechloride-methanol-conc. ammonia, 90:9:1.5. By evaporating the combinedfractions with Rf 0.480 there was obtained chromatographicallyhomogeneous3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-azithromycin3,6-hemiketal with the following physical-chemical constants:

TLC Ethyl acetate-triethyl amine, 95:5 Rf 0.447

IR(CDCl₃) cm⁻¹: 3468, 2976, 1713, 1638, 1459, 1382, 1197, 1116, 1068,1049, 1014, 963.

¹H NMR (300 MHz,CDCl₃) δ: 4.94 (H-13), 4.21 (H-¹′), 3.74 (H-5), 3.51(H-5′), 3.23 (H-2′), 2.57 (H-2), 2.49 (H-3′), 2.23 /3′N(CH₃)₂/, 2.06(H-4), 1.74 (H-8), 1.67 (H-4′a), 1.39 (6-CH₃), 1.28 (2-CH₃), 1.25(H-4′b), 1.22 (5′-CH₃), 1.23, (4-CH₃), 1.10 (10-CH₃), 1.04 (8-CH₃), 0.92(14-CH₃).

¹³C NMR (75 MHz, CDCl₃) δ: 176.9 (C-1), 106.1 (C-1′), 102.3 (C-3), 94.8(C-5), 82.4 (C-6), 69.7 (C-2′), 68.5 (C-11), 66.4 (C-9), 65.3 (C-3′),61.6 (C-10), 49.3 (C-2), 41.6 (C-7), 40.1 /3′N(CH₃)₂/, 31.0 (C-8), 28.2(C-4′), 26.4 (6-CH₃), 20.8 (5′-CH₃), 13.6 (2-CH₃), 12.6 (4-CH₃), 11.4(14-CH₃).

FAB-MS m/z 589

EXAMPLE 183-De(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-azithromycin3,6-hemiketal 11,12-cyclic carbonate

To a solution of3-de(2,6-dideoxy-3-C-methyl-3-O-methyl-α-L-ribohexopyranosyl-oxy)-azithromycin3,6-hemiketal (1 g) from Example 17 in ethyl acetate (30 ml), ethylenecarbonate (0.5 g) and potassium carbonate (0.5 g) were added. Thereaction suspension was stirred under reflux for 10 hours, left standingfor 16 hours at room temperature and then filtered. Ethyl acetate wasrinsed with saturated NaCl solution and water, dried over CaCl₂,filtered and evaporated, yielding 1.05 g of an oily residue. Afterseparation on a silica gel column using the system methylenechloride-methanol-conc. ammonia, 90:9:0.5, there was obtainedchromatographically homogenous title product with the followingphysical-chemical constants:

TLC Ethyl acetate-tniethyl amine, 95:5 Rf 0.514

IR(CDCl₃) cm⁻¹: 3498, 2975, 2941, 1812, 1724, 1638, 1459, 1381, 1359,1333, 1292, 1234, 1173, 1115, 1082, 1045, 1015, 966.

¹H NMR (300 MHz, CDCl₃) δ: 5.03 (H-13), 4.61 (H-11), 4.23 (H-1′), 3.73(H-5), 3.52 (H-5′), 3.25 (H-2′), 3.18 (H-9a), 2.90 (H-10), 2.54 (H-2),2.50 (H-3′), 2.28 /3′N(CH₃)₂/, 2.10 (H-4), 2.07 (9a-NCH₃), 1.76 (H-7a),1.95 (H-8), 1.86 (H-14a), 1.67 (H-4′a), 1.57 (H-9b), 1.55 (H-14b), 1.45(12-CH₃), 1.37 (6-CH₃), 1.30 (2-CH₃), 1.28 (H-4′b), 1.23 (5′-CH₃), 1.24(4-CH₃), 1.13 (H-7b), 1.18 (10-CH₃), 0.90 (8-CH₃), 0.92 (14-CH₃).

¹³C NMR (75 MHz, CDCl₃) δ: 176.1 (C-1), 153.5 C═O carbonate), 106.1(C-1′), 101.6 (C-3), 93.6 (C-5), 83.7 (C-12), 82.7 (C-6), 78.9 (C-11),77.9 (C-13), 69.6 (C-2′), 69.4 (C-5′), 63.6 (C-9), 65.3 (C-3′), 60.1(C-10), 49.9 (C-2), 46.6 (C-4), 41.8 (C-7), 40.0/3′N(CH₃)₂/, 33.4(9a-CH₃), 28.0 (C-4′), 26.8 (C-8), 25.1 (6-CH₃), 22.3 (C-14), 20.8(5′-CH₃), 19.4 (8-CH₃), 14.1 (12-CH₃), 13.9 (2-CH₃), 12.1 (4-CH₃), 12.9(10-CH₃), 10.1 (14-CH₃).

What is claimed is:
 1. A compound of the formula (I)

characterized in that R¹ individually stands for hydroxyl or aL-cladinosyl group of the formula (II)

wherein R² individually stands for a silyl group, R³ individually standsfor hydrogen or together with R⁶ stands for an ether group, R⁴individually stands for hydrogen, (C₁-C₄)acyl group or —COO—(CH₂)_(n)—Argroup, wherein n is 1-7 and Ar individually stands for an unsubstitutedor substituted aryl group with up to 18 carbon atoms, R⁵ individuallystands for hydrogen, methyl group or —COO—(CH₂)_(n)—Ar group, wherein nis 1-7 and Ar individually stands for an unsubstituted or substitutedaryl group with up to 18 carbon atoms, R⁶ individually stands forhydroxyl group or together with R³ stand for an ether group, R⁷individually stands for hydrogen, (C₁-C₁₂)alkyl group, silyl group ortogether with R⁸ and C-11/C-12 carbon atoms stands for a cycliccarbonate, R⁸ individually stands for hydrogen, (C₁-C₁₂)alkyl group,silyl group or together with R⁷ and C-11/C-12 carbon atoms for a cycliccarbonate, or a pharmaceutically acceptable addition salts with aninorganic or organic acid.
 2. Compound according to claim 1,characterized in that R¹ stands for L-cladinosyl group, R² and R⁷ aremutually the same and stand for trimethylsilyl group, R³ and R⁸ aremutually the same and stand for hydrogen, R⁴ and R⁵ are mutually thesame and stand for benzyloxycarbonyl group and R⁶ is hydroxyl group. 3.Compound according to claim 1, characterized in that R¹ stands forL-cladinosyl group, R² stands for trimethylsilyl group, R³, R⁷ and R⁸are mutually the same and stand for hydrogen, R⁴ and R⁵ are mutually thesame and stand for benzyloxy-carbonyl group and R₆ is hydroxyl group. 4.Compound according to claim 1, characterized in that R¹ stands forL-cladinosyl group, R² and R⁷ are mutually the same and stand fortrimethylsilyl group, R³ stands for hydrogen, R⁴ and R⁵ are mutually thesame and stand for benzyloxy-carbonyl group, R⁶ is hydroxyl group and R⁸is methyl.
 5. Compound according to claim 1, characterized in that R¹stands for L-cladinosyl group, R² stands for trimethylsilyl group, R³and R⁸ are mutually the same and stand for hydrogen, R⁴ and R⁵ aremutually the same and stand for benzyloxy-carbonyl group, R⁶ stands forhydroxyl group and R⁷ is methyl.
 6. Compound according to claim 1,characterized in that R¹ stands for L-cladinosyl group, R² stands fortrimethylsilyl group, R³ and R⁷ are mutually the same and stand forhydrogen, R⁴ and R⁵ are mutually the same and stand forbenzyloxy-carbonyl group, R⁶ is hydroxyl group and R⁸ is methyl. 7.Compound according to claim 1, characterized in that R¹ and R⁶ aremutually the same and stand for hydroxyl group, R³, R⁴ and R⁸ aremutually the same and stand for hydrogen and R⁵ and R⁷ are mutually thesame and stand for methyl.
 8. Compound according to claim 1,characterized in that R¹ and R⁶ are mutually the same and stand forhydroxyl group, R³, R⁴ and R⁷ are mutually the same and stand forhydrogen and R⁵ and R⁸ are mutually the same and stand for methyl. 9.Compound according to claim 1, characterized in that R¹ and R⁶ aremutually the same and stand for hydroxyl group, R³, R⁷ and R⁸ aremutually the same and stand for hydrogen, R⁴ is acetyl and R⁵ is methyl.10. Compound according to claim 1, characterized in that, R¹ and R⁶ aremutually the same and stand for hydroxyl group, R³ and R⁸ are mutuallythe same and stand for hydrogen, R⁴ is acetyl and R⁵ and R⁷ are mutuallythe same and stand for methyl.
 11. Compound according to claim 1,characterized in that R¹ and R⁶ are mutually the same and stand forhydroxyl group, R³ and R⁷ are mutually the same and stand for hydrogen,R⁴ is acetyl and R⁵ and R⁸ are mutually the same and stand for methyl.12. Compound according to claim 1, characterized in that R¹ stands forhydroxyl group, R³ together with R⁶ stands for an ether group, R⁴ isacetyl, R⁵ is methyl, R⁷ and R⁸ are mutually the same and stand forhydrogen.
 13. Compound according to claim 1, characterized in that R¹stands for hydroxyl group, R³ together with R⁶ stands for an ethergroup, R⁴ is acetyl, R⁵ and R⁷ are mutually the same and stand formethyl, and R⁸ is hydrogen.
 14. Compound according to claim 1,characterized in that R¹ stands for hydroxyl group, R³ together with R⁶stands for an ether group, R⁴ is acetyl, R⁵ and R⁸ are mutually the sameand stand for methyl and R⁷ is hydrogen.
 15. Compound according to claim1, characterized in that R¹ stands for hydroxyl group, R³ together withR⁶ stands for an ether group, R⁴, R⁷ and R⁸ are mutually the same andstand for hydrogen, and R⁵ is methyl.
 16. Compound according to claim 1,characterized in that R¹ stands for hydroxyl group, R³ together with R⁶stands for an ether group, R⁴ and R⁸ are mutually the same and stand forhydrogen and R⁵ and R⁷ are mutually the same and stand for methyl. 17.Compound according to claim 1, characterized in that R¹ stands forhydroxyl group, R³ together with R⁶ stands for an ether group, R⁴ and R⁷are mutually the same and stand for hydrogen and R⁵ and R⁸ are mutuallythe same and stand for methyl.
 18. Compound according to claim 1,characterized in that R¹ stands for hydroxyl group, R³ together with R⁶stands for an ether group, R⁴ is hydrogen, R⁵ is methyl and R⁷ and R⁸together with C-11/C-12 carbon atoms stand for a cyclic carbonate.
 19. Aprocess for the preparation of compounds of the formula (I)

wherein R¹ individually stands for hydroxyl or a L-cladinosyl group ofthe formula (II)

wherein R² individually stands for a silyl group, R³ individually standsfor hydrogen or together with R⁶ stands for an ether group, R⁴individually stands for hydrogen, (C₁-C₄)acyl group or —COO—(CH₂)_(n)—Argroup, wherein n is 1-7 and Ar individually stands for unsubstituted orsubstituted aryl group with up to 18 carbon atoms, R⁵ individuallystands for hydrogen, methyl group or —COO—(CH₂)_(n)—Ar group, wherein nis 1-7 and Ar individually stands for unsubstituted or substituted arylgroup with up to 18 carbon atoms. R⁶ individually stands for hydroxylgroup or together with R³ stands for an ether group, R⁷ individuallystands for hydrogen, (C₁-C₁₂)alkyl group, silyl group or together withR⁸ and C-11/C-12 carbon atoms stands for acyclic carbonate, R⁸individually stands for hydrogen, (C₁-C₁₂)alkyl group, silyl group ortogether with R⁷ and C-11/C-12 carbon atoms stands for a cycliccarbonate, or a pharmaceutically acceptable addition salt with aninorganic or organic acid, characterized in that (I) azithromyein of theformula (I), wherein R¹ stands for L-cladinosyl group of the formula(II), R² stands for a silyl group, and wherein, R³, R⁴, R⁷ and R⁸ aremutually the same and stand for hydrogen, R⁵ is methyl and R⁶ ishydroxyl group, is subjected to a reaction with organic carboxylic acidchlorides of the formula (III) ClCOO(CH₂)_(n)—Ar  (III) wherein n is 1-7and Ar individually stands for unsubstituted or substituted aryl groupwith up to 18 carbon atoms in the presence of a first bases in a firstreaction-inert solvent, yielding a compound of the general formula (I),wherein R¹ stands for L-cladinosyl group of formula (II), R² stands fora silyl group, and wherein, R³, R⁷ and R⁸ are mutually the same andstand for hydrogen, R⁴ and R⁵ are mutually the same and stand forbenzyloxycarbonyl group and R⁶ is hydroxyl group, which is subsequentlysubjected to a selective silylation of hydroxyl groups in A/4 4″- and11-positions with 2-5 equimolar excess of a silylating agent in anorganic inert solvent at a temperature 0-5° C. during 5-8 hours,yielding a compound of the formula (I), wherein R¹ stands fortrimethylsilyl group, R³ and R⁸ are mutually the same and stand forhydrogen, R⁴ and R⁵ are mutually the same and stand forbenzyloxycarbonyl group and R⁶ is hydroxyl group, or in B/ 4″-positionwith 1.1-2 equimolar excess of a silylating agent, in an organic inertsolvent at a temperature 0.5° C., during a 1 hour, yielding a compoundof formula (I), wherein R¹ stands for L-cladinosyl group of the formula(II), R² stands for trimethylsilyl group, R³, R⁷ and R⁸ are mutually thesame and stand for hydrogen, R⁴ and R⁵ are mutually the same and standfor benzyloxycarbonyl group and R⁶ stands for hydroxyl group, which arethen subjected to O-alkylation with 1.3 to 10 molar excess ofcorresponding alkylating agent in the presence of 1.1-8.5 moles of asecond base in a second reaction-inert solvent at an O-alkylationtemperature from −15° C. to room temperature yielding in the case of A/a compound of formula (I), wherein R¹ stands for L-cladinosyl group ofthe formula III), R² and R⁷ are mutually the same and stand fortrimethylsilyl group, R³ stands for hydrogen, R⁴ and R⁵ are mutually thesame and stand for benzyloxycarbonyl group, R⁶ is hydroxyl group and R⁸is methyl, or in the case of B/ a mixture of a compound of the formula(I), wherein R¹ stands for L-cladinosyl group of the formula (II), R²stands for trimethylsilyl group, R³ and R⁸ are mutually the same andstand for hydrogen, R⁴ and R⁵ are mutually the same and stand forbenzyloxycarbonyl group, R⁶ stands for hydroxyl group and R⁷ is methyl,and of a compound of the general formula (I), wherein R¹ stands forL-cladinosyl group of the formula (II), R³ ad R⁷ are mutually the sameand stand for hydrogen, R⁴ and R⁵ are mutually the same and stand forbenzyloxycarbonyl group, R⁶ is hydroxyl group and R⁸ is methyl, whichare then subjected to deprotection of the protecting groups in 2′- and3′-positions in a solution of a first lower alcohol in the presence ofNaOAc/HOAc buffer (pH 5) and of a catalyst in hydrogen atmosphere at apressure of 1-20 bars and then after isolation, to desilylation in 4″-and 11-positions in a second lower alcohol in the presence of formicacid, yielding in the case of A/ a compound of the general formula (I),wherein R¹ stands for L-cladinosyl group of the formula (II), R¹, R²stands for a silyl group, and wherein, R³, R⁴, R⁵ and R⁷ are mutuallythe same and stand for hydrogen, R⁶ is hydroxyl group and R⁸ is methyl,or in the case of B/ a mixture of a compound of the general formula (I),wherein R¹ stands for L-cladinosyl group of the formula (II), R², R³,R⁴, R⁵ and R⁸ are mutually the same and stand for hydrogen, R⁶ishydroxyl group and R⁷ is methyl, and of a compound of formula (I),wherein R¹, R² stands for a silyl group, and wherein, R³, R⁴, R⁵, R⁷ andR⁸ have the meanings as given for deprotection in the case of A/, whichare then subjected to reductive 3′-N-methylation with 1-3 equivalents offormaldehyde (37%) in the presence of an equal or double quantity offormic acid (98-100%) and hydrogenation catalyst or of some otherhydrogen source, in a third reaction-inert solvent at an elevatedtemperature, yielding in the case of A/ a compound of the generalformula (I), wherein R¹ stands for L-cladinosyl group of the formula(II), R² stands for a silyl group, and wherein, R³, R⁴ and R⁷ aremutually the same and stand for hydrogen, R⁵ and R⁸ are mutually thesame and stand for methyl and R⁶is hydroxyl group, or in the case of B/a mixture of a compound of the general formula (I), wherein R¹ standsfor L-cladinosyl group of the formula (II), R² stands for a silyl group,and wherein, R³, R⁴, R⁵ and R⁸ are mutually the same and stand forhydrogen R⁵ and R⁷ are mutually the same and stand for methyl and R⁶ ishydroxyl group and R⁷ is methyl, and of a compound of formula (I),wherein R¹, R², R³, R⁴, R⁵, R⁷ and R⁸ have the meanings as given fordeprotection in the case of A/, which is then optionally subjected toseparation on a silica gel column, yielding a chromatographicallyhomogeneous compound of formula (I), wherein R¹ stands for L-cladinosylgroup, R², R³, R⁴, and R⁸ are mutually the same and stand for hydrogen,R⁵ and R⁷ are mutually the same and stand for methyl and R⁶ stands forhydroxyl group (11-O-methyl-azithromycin) and a compound of formula (I),wherein R¹, R², R³, R⁴, R⁵ R⁶, R⁷ and R⁸ have the meanings as given for3′-N-methylation in the case of A/ (12-O-methyl azithromycin), or that(II) acithromycin or its 11-O-methyl- and 12-O-methyl-derivativesobtained according to the process (I) are optionally subjected to areaction of hydrolysis with diluted inorganic acids yielding a compoundof formula (I), wherein R¹ and R⁶ are mutually the same and stand forhydroxyl group, R³, R⁴ and R⁸ are mutually the same and stand forhydrogen, and R⁵ and R⁷ are mutually the same and stand for methyl, or acompound of formula (I), wherein R¹ and R⁶ are mutually the same andstand for hydroxyl group, R³, R⁴ and R⁷ are mutually the same and standfor hydrogen, and R⁵ and R⁸ are mutually the same and stand for methyl,which are then optionally subjected to a reaction of selective acylationof hydroxyl group in 2′-position with chlorides and anhydrides ofcarboxylic acids with up to 4 carbon atoms in a fourth reaction-inertorganic solvent yielding a compound of formula (I), wherein R¹ and R⁶are mutually the same and stand for hydroxyl group, R³, R⁷ and R⁸ aremutually the same and stand for hydrogen, R⁴ is acetyl and R⁵ is methyl,or a compound of formula (I), wherein R¹ and R⁶ are mutually the sameand stand for hydroxyl group, R³ and R⁸ are mutually the same and standfor hydrogen, R⁴ is acetyl and R⁵ and R⁷ are mutually the same and standfor is methyl, or a compound of formula (I), wherein R¹ and R⁶ aremutually the same and stand for hydroxyl group, R³ and R⁷ are mutuallythe same and stand for hydrogen, R⁴ is acetyl and R⁵ and R⁸ are mutuallythe same and stand for methyl, which are then optionally subjected tooxidation with Jones reagent or according to a modified Moffat-Pfitznerprocess in the presence of dimethyl sulfoxide and pyridiniumtrifluoracetate as a catalyst, in a fourth reaction-inert organicsolvent at a temperature from 10° C. to room temperature, yielding acompound of formula (I), wherein R¹ stands for hydroxyl group, R³together with R⁶ stands for an ether group, R⁴ is acetyl, R⁵ is methyl,R⁷ and R⁸ are mutually the same and stand for hydrogen, or a compound offormula (I), wherein R¹ stands for a hydroxyl group, R³ together with R⁶stands for an ether group, R⁴ is acetyl, R⁵ and R⁷ are mutually the sameand stand for methyl and R⁸ is hydrogen, or a compound of the generalformula (I), wherein R¹ stands for a hydroxyl group, R³ together with R⁶stands for an ether group, R⁴is acetyl, R⁵ and R⁸ are mutually the sameand stand for methyl and R⁷ is hydrogen, which are then subjected to adeacylation reaction in 2′-position by means of solvolysis in a thirdlower alcohol at room temperature, yielding a compound of formula (I),wherein R¹ stands for hydroxyl group, R³ together with R⁶ stands for anether group, R⁴, R⁷ and R⁸ are mutually the same and stand for hydrogen,and R⁵ is methyl, or a compound of formula (I), wherein R¹ stands forhydroxyl group, R³ together with R⁶ stands for an ether group, R⁴ and R⁸are mutually the same and stand for hydrogen, and R⁵ and R⁷ are mutuallythe same and stand for hydroxyl group, R³ together with R⁶ stands for anether group, R⁴ and R⁷ are mutually the same and stand for hydrogen, andR⁵ and R⁸ are mutually the same and stand for methyl, and then acompound of formula (I), wherein R¹ stands for hydroxyl group, R³together with R⁶ stands for an ether group, R⁴, R⁷ and R⁸ are mutuallythe same and stand for hydrogen and R⁵ is methyl, are optionallysubjected to a reaction with ethylene carbonate in the presence of aninorganic or organic in a fifth reaction-inert solvent yielding acompound of formula (I), wherein R¹ stands for hydroxyl group, R³together with R⁶ stands for an ether group, R⁴ is hydrogen, R⁵ ismethyl, and R⁷ and R⁸ together with C-11/C-12 carbon atoms stand for acyclic carbonate.
 20. Pharmaceutical composition useful in the treatmentof bacterial infections in humans or animals comprising antibacteriallyeffective amounts of a compound of formula (I) or its pharmaceuticallyacceptable addition salts according to claim 1 in a combination with apharmaceutically acceptable carrier.
 21. A method of treatment ofbacterial infections in humans or animals comprising the administrationof an antibacterially effective amount of a compound of formula (I) orits pharmaceutically acceptable salts with inorganic or organic acidsaccording to claim 1 in a combination with a pharmaceutically acceptablecarrier to humans or animals in need of such treatment.